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Transfection Calculator

Scale transfection recipes across vessel formats, convert PEI N:P ratios, and split AAV co-transfection mixes—no account. Built-in AI agent assistant support.

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Key facts

Key facts about Transfection Calculator
FactValue
InputsTransfection mode (standard/co-transfection), reagent, vessel format, DNA mass, stock conc., ratio mode (N:P or mass), replicates, overage %
Reagents supportedPEI MAX, PEI linear 25 kDa, PEI branched 25 kDa, Lipofectamine 2000, Lipofectamine 3000, FuGENE HD, CaPO₄
Vessel formats96W (0.32 cm²), 24W (1.9 cm²), 12W (3.8 cm²), 6W (9.6 cm² baseline), T25 (25 cm²), T75 (75 cm²), D100 (55 cm²), D150 (152 cm²), custom
Co-transfection presetsAAV triple-transfection (pHelper, pRC, Transfer), lentiviral 4-plasmid system, custom multi-plasmid split
Complex protocol2-Tube Opti-MEM split (Tube A: DNA + Media 50%, Tube B: Reagent + Media 50%)
Master mix scaling1–384 replicates with 0–25% customizable overage (default 10%)
Runs in browserYes — calculations run locally
Account requiredNo

What it does

Scaling a 6-well PEI recipe to a T-75 flask or splitting AAV triple-transfection plasmid masses leads to manual calculation errors, wrong N:P ratios, and wasted transfection reagents. TransfectionWise turns your target vessel, reagent selection, stock concentrations, and plasmid ratios into a complete 2-tube transfection protocol with exact pipetting microliters—updated live as you type.

Select standard single-plasmid or multi-plasmid co-transfection mode, then choose your reagent: PEI MAX, PEI linear 25 kDa, PEI branched 25 kDa, Lipofectamine 2000, Lipofectamine 3000, FuGENE HD, or calcium phosphate. Pick from eight standard vessel formats (96-well through 15 cm dish) or enter a custom surface area in cm². PEI reagents automatically toggle between N:P molar charge ratio and mass ratio; Lipofectamine 3000 calculates optional P3000 enhancer volume (2.0 µL per µg DNA). For co-transfections, load AAV or lentiviral presets to split total DNA by molar ratio using plasmid base-pair lengths.

Results display DNA per well (µg), stock DNA pipetting volume (µL), reagent volume (µL), and a 50/50 2-tube Opti-MEM media split (Tube A: DNA + Media, Tube B: Reagent + Media) with incubation timing. Adjust replicate count (1–384) and master-mix overage (0–25%) for bulk prep. Copy tab-separated data, download CSV files, print a checkbox bench sheet, or save named recipes in local browser storage.

Why researchers use it

  • Scale transfection recipes across vessel formats by surface area ratio
  • Convert PEI N:P charge ratios to reagent mass ratios instantly
  • Split AAV and lentiviral plasmid mixes by base pair-weighted molar ratio
  • Get 2-tube complexation breakdown for Tube A and Tube B diluent media
  • Calculate master-mix totals with customizable overage percentage
  • Store cell-line protocols locally in browser storage without an account

Best for

  • Scaling PEI MAX or Lipofectamine 3000 from 6-well plates to T-75 flasks
  • Preparing AAV triple-transfection or lentiviral 4-plasmid packaging mixes
  • Optimizing PEI N:P charge ratios for transient protein expression
  • Generating 2-tube complexation protocols for multi-well plate screens
  • Standardizing transfection protocols across team members without login

When to use this vs alternatives

Choose TransfectionWise when you need reagent-agnostic scaling, PEI N:P conversion, AAV/lentiviral molar plasmid splits, or 2-tube complexation protocols in one browser session. Use the Cell Seeding Density Calculator prior to complex addition to verify cell confluency and seeding counts. Vendor calculators and spreadsheets remain useful for single static recipes, but lack multi-reagent flexibility and 2-tube bench exports.

What makes it different

Vendor calculators restrict you to a single proprietary reagent catalog, while generic spreadsheets break when switching between PEI N:P ratios and Lipofectamine mass ratios or when adjusting multi-plasmid base-pair stoichiometry.

TransfectionWise provides a reagent-agnostic workspace with automatic vessel surface-area scaling, real-time N:P charge ratio conversion, base pair-weighted co-transfection plasmid splitting, 2-tube Opti-MEM protocol generation, CSV/print export, and browser-local protocol storage—all on one page without server uploads.

How to get started

  1. Open the Calculator tab and select Standard or Co-transfection mode.
  2. Select Reagent (PEI, Lipofectamine, FuGENE HD, or CaPO₄) and Vessel format.
  3. Enter DNA mass, plasmid stock concentration, and N:P ratio or reagent mass ratio.
  4. For multi-plasmid runs, open Co-transfection presets to load AAV or lentiviral ratios, or enter plasmid base-pair sizes.
  5. Review live results: stock pipetting µL, 2-tube media split, and master-mix totals with overage.
  6. Click Copy, CSV download, or Print Bench Sheet—or click Save protocol to store the recipe locally.

Frequently asked questions

How is reagent volume calculated for PEI transfection?
PEI volume is calculated from the N:P molar charge ratio, which represents protonatable nitrogen in PEI relative to phosphate in DNA. The calculator converts N:P to a mass ratio using a monomer formula weight ratio (43.07/330). For stock concentrations in mg/mL or mM, microliters are derived instantly and flagged if outside recommended ranges (such as N:P 15–23 for PEI MAX).
How does vessel surface area scaling work across plate formats?
All DNA mass and reagent volumes scale linearly by surface area ratio relative to a 6-well plate baseline (9.6 cm²). For example, scaling to a 24-well plate (1.9 cm²) applies a 0.198× multiplier, while a T-75 flask (75 cm²) applies a 7.81× multiplier. Custom surface area inputs scale with the same linear ratio.
How does co-transfection molar-to-mass conversion work for AAV and lentivirus?
In co-transfection mode, total DNA mass is distributed across plasmids based on entered molar ratios and plasmid lengths in base pairs (bp). Mass fraction for plasmid i is calculated as (molar_ratio_i × bp_i) ÷ Σ(molar_ratio_j × bp_j). This ensures stoichiometric equimolar packaging for AAV triple-transfection and 4-plasmid lentiviral systems.
Why is a 2-tube complex preparation protocol required?
Proper transfection complex formation requires separate initial dilution of DNA and cationic reagent in serum-free medium (such as Opti-MEM) before combining. TransfectionWise automatically splits total diluent volume 50/50 into Tube A (DNA + Media) and Tube B (Reagent + Media), preventing premature charge neutralization or reagent precipitation.
Does Lipofectamine 3000 include P3000 enhancer volume?
Yes. When Lipofectamine 3000 is selected for DNA transfection, P3000 enhancer volume is automatically included at 2.0 µL per µg of plasmid DNA. You can toggle P3000 off when performing siRNA transfections or using custom reagent ratios.
Can I use ChatGPT, Cursor, or another AI agent with TransfectionWise?
Yes. The workspace includes an in-browser AI assistant that helps validate inputs, explain N:P charge ratios, and interpret complexation warnings. External AI agents and automated scripts can also run transfection calculations via [API & MCP](/tools/developers) using the tool identifier pepkio_transfectionwise.

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Last updated . Pepkio builds free lab calculators alongside bioinformatics CRO services.